Metabolomics analysis reveals changes related to pseudocyst formation induced by iron depletion in Trichomonas vaginalis.

BACKGROUND: Iron is an essential element for cellular functions, such as energy metabolism. Trichomonas vaginalis, a human urogenital tract pathogen, is capable of surviving in the environment without sufficient iron supplementation. Pseudocysts (cyst-like structures) are an environmentally tolerated stage of this parasite while encountering undesired conditions, including iron deficiency. We previously demonstrated that iron deficiency induces more active glycolysis but a drastic downregulation of hydrogenosomal energy metabolic enzymes. Therefore, the metabolic direction of the end product of glycolysis is still controversial. METHODS: In the present work, we conducted an LC‒MS-based metabolomics analysis to obtain accurate insights into the enzymatic events of T. vaginalis under iron-depleted (ID) conditions. RESULTS: First, we showed the possible digestion of glycogen, cellulose polymerization, and accumulation of raffinose family oligosaccharides (RFOs). Second, a medium-chain fatty acid (MCFA), capric acid, was elevated, whereas most detected C18 fatty acids were reduced significantly. Third, amino acids were mostly reduced, especially alanine, glutamate, and serine. Thirty-three dipeptides showed significant accumulation in ID cells, which was probably associated with the decrease in amino acids. Our results indicated that glycogen was metabolized as the carbon source, and the structural component cellulose was synthesized at same time. The decrease in C18 fatty acids implied possible incorporation in the membranous compartment for pseudocyst formation. The decrease in amino acids accompanied by an increase in dipeptides implied incomplete proteolysis. These enzymatic reactions (alanine dehydrogenase, glutamate dehydrogenase, and threonine dehydratase) were likely involved in ammonia release. CONCLUSION: These findings highlighted the possible glycogen utilization, cellulose biosynthesis, and fatty acid incorporation in pseudocyst formation as well as NO precursor ammonia production induced by iron-depleted stress.

Serious bacterial infections in young children with fever without source after discharge from emergency department: A National Health Insurance database cohort study.

BACKGROUND: Well-appearing febrile young children discharged from the emergency department (ED) after medical assessment are still at risk for serious bacterial infections (SBI). The incidence of SBI and the effectiveness of laboratory tests in the pneumococcal conjugate vaccine era remain unknown. METHODS: We conducted a study using Taiwan's National Health Insurance claims data from 2004 to 2014. Children aged 2-24 months discharged from the ED with a diagnosis compatible with fever without source (FWS) were enrolled. RESULTS: The study identified 431,884 children from the ED with FWS. 13.53% of the children had revisits, 8.62% needed hospitalization and 1.57% developed SBI. Younger children had a higher SBI rate, but a lower revisit rate. The revisit rate was 12.22% for children aged 2-6 months, 13.61% for children aged 7-12 months and 13.77% for children aged 13-24 months (p < 0.0001). The SBI rate was 4.44% for children aged 2-6 months, 1.85% for children aged 2-6 months and 0.96% for children aged 13-24 months (p < 0.0001). Children with hemogram tests, compared to those without, had a higher revisit rate (16.30% vs. 13.15%, p < 0.0001), and a higher SBI rate in the children aged 13-24 months (1.30% vs. 0.92%, p < 0.0001); furthermore, children with urinalysis had a significantly higher revisit rate (14.42% vs. 13.24%, p < 0.0001) and higher SBI rate (2.10% vs. 1.40%, p < 0.0001). CONCLUSION: Children with FWS aged 2-24 months who were discharged from ED after blood test and urinalysis were still at risk for SBI, especially those aged 2-6 months.

Immunogenicity and Safety of AS03-adjuvanted H5N1 Influenza Vaccine in Children 6-35 Months of Age: Results From a Phase 2, Randomized, Observer-blind, Multicenter, Dose-ranging Study.

BACKGROUND: This phase 2 observer-blind, randomized, multicenter, dose-ranging study evaluated immunogenicity and safety of different formulations of an AS03-adjuvanted H5N1 influenza vaccine in children 6-35 months of age. METHODS: One hundred eighty-five children randomized into 5 groups [1.9 µg hemagglutinin (HA)/AS03B, 0.9 µg HA/AS03C, 1.9 µg HA/AS03C, 3.75 µg HA/AS03C or 3.75 µg HA/AS03D] were to receive 2 doses administered 21 days apart (primary vaccination). AS03 was classified by amount of DL-α-tocopherol, with AS03B the highest amount. One year later, all subjects were to receive unadjuvanted 3.75 µg HA as antigen challenge. Immunogenicity was assessed 21 days after primary vaccination (day 42) and 7 days after antigen challenge (day 392). Immunogenicity-fever index, based on hemagglutination inhibition and microneutralization antibody titers at day 42 and fever 7 days after each vaccination, was used to guide the selection of an acceptable formulation. RESULTS: After primary vaccination, formulations elicited strong homologous immune responses with all subjects' hemagglutination inhibition titers ≥1:40 post-vaccination. Immunogenicity-fever index based on hemagglutination inhibition and microneutralization assays showed that 1.9 µg HA/AS03B ranked the highest. Antibody levels persisted >4 times above baseline 12 months after primary vaccination with all formulations (day 385). Antibodies increased >4-fold after antigen challenge (day 392/day 385) with 1.9 µg HA/AS03B, 0.9 µg HA/AS03C and 1.9 µg HA/AS03C formulations. Overall per subject, the incidence of fever ranged from 28.6% (3.75 µg HA/AS03D) to 60.5% (1.9 µg HA/AS03B). CONCLUSIONS: All formulations were highly immunogenic and demonstrated acceptable safety profiles, with the 1.9 µg HA/AS03B providing the most favorable balance of immunogenicity versus reactogenicity for use in children 6-35 months of age.

Effect of Phosphatase Activity of the Control of Virulence Sensor (CovS) on Clindamycin-Mediated Streptolysin O Production in Group A Streptococcus.

Severe manifestations of group A Streptococcus (GAS) infections are associated with massive tissue destruction and high mortality. Clindamycin (CLI), a bacterial protein synthesis inhibitor, is recommended for treating patients with severe invasive GAS infection. Nonetheless, the subinhibitory concentration of CLI induces the production of GAS virulent exoproteins, such as streptolysin O (SLO) and NADase, which would enhance bacterial virulence and invasiveness. A better understanding of the molecular mechanism of how CLI triggers GAS virulence factor expression will be critical to develop appropriate therapeutic approaches. The present study shows that CLI activates SLO and NADase expressions in the emm1-type CLI-susceptible wild-type strain but not in covS or control of virulence sensor (CovS) phosphatase-inactivated mutants. Supplementation with Mg2+, which is a CovS phosphatase inhibitor, inhibits the CLI-mediated SLO upregulation in a dose-dependent manner in CLI-susceptible and CLI-resistant strains. These results not only reveal that the phosphorylation of response regulator CovR is essential for responding to CLI stimuli, but also suggest that inhibiting the phosphatase activity of CovS could be a potential strategy for the treatment of invasive GAS infection with CLI.

Probiotic Lactobacillus casei: Effective for Managing Childhood Diarrhea by Altering Gut Microbiota and Attenuating Fecal Inflammatory Markers.

BACKGROUND: Acute diarrhea is a major cause of childhood morbidity and an economic burden for families. The aim of this study is to explore the effect of probiotics on clinical symptoms, intestinal microbiota, and inflammatory markers during childhood diarrhea. METHODS: Children (n = 81) aged six months to six years (mean age 2.31 years) hospitalized for acute diarrhea were randomized to receive probiotics (Lactobacillus casei variety rhamnosus; n = 42) or no probiotics (n = 39) orally twice daily for seven days. Feces samples were also collected to evaluate microbial content using a traditional agar plate and next-generation sequencing. Immunoglobulin A (IgA), lactoferrin, and calprotectin were determined by enzyme-linked immunosorbent assay (ELISA) and compared in different groups. Other clinical symptoms or signs, including fever, vomiting, diarrhea, abdominal pain, bloated abdomen, daily intake, appetite, and body weight were also assessed. RESULTS: Data were collected from 81 individuals across three different time points. Total fecal IgA levels in fecal extracts of the probiotics group were higher than those in the control group, reaching statistical significance (p < 0.05). Concentrations of fecal lactoferrin and calprotectin were significantly downregulated in patients with probiotic Lactobacillus casei variety rhamnosus (Lc) consumption compared to those of the control (p < 0.05). Probiotic Lc administration may be beneficial for gut-microbiota modulation, as shown by the data collected at one week after enrollment. Counts of Bifidobacteria and Lactobacillus species were elevated in stool culture of the probiotic group. Appetite and oral intake, body-weight gain, abdominal pain, bloating, as well as bowel habits (diarrhea) were much better in children receiving probiotics compared with those in the control group. CONCLUSION: Fecal IgA increased during acute diarrhea under Lc treatment; in contrast, fecal lactoferrin and calprotectin were downregulated during acute diarrhea under Lc treatment. Probiotic Lc may be a useful supplement for application in children during acute diarrhea to reduce clinical severity and intestinal inflammatory reaction.

Risk Factors and Outcome Analysis in Children with Carbon Monoxide Poisoning.

BACKGROUND: Carbon monoxide (CO) poisoning is one of the common causes of poisoning in patients and can result in significant morbidity and mortality. However, few studies have focused on the pediatric group. METHODS: We retrospectively reviewed children (age < 18 years) with CO poisoning from nonfire accidents at a tertiary medical center in Taiwan from 2002 to 2010. We analyzed the patients' characteristics, management, and outcome; compared the data of patients who received hyperbaric oxygen (HBO) to those who received normobaric oxygen (NBO) therapy; and identified the ri0sk factors for patients who developed delayed neurological sequelae (DNS) or permanent neurological sequelae (PNS). RESULTS: A total of 81 children were enrolled. The annual case number increased from five cases in 2002 to 20 in 2010, particularly during the cold months (December to February). The most common source of exposure was an indoor heating system (54.3%). The most common presenting symptoms were vomiting (32.1%) and consciousness changes (30.9%). HBO treatment tended to be administered to patients with a higher initial COHb (%) (p < 0.001), an initial Glasgow coma scale change (p < 0.001), and admission to the hospital (p = 0.002). After multivariate analysis, treatment in the intensive care unit because of prolonged loss of consciousness (p = 0.002) was the only independent risk factor for patients with DNS; only rescue by a ventilator (p < 0.001) was an independent risk factor for patients with PNS. In comparison to the NBO therapy, HBO treatment did not show benefit or harm to patients according to the incidence of inducing DNS or PNS after multivariate analysis. CONCLUSION: For those with treatment in the intensive care unit because of prolonged loss of consciousness and rescue by a ventilator, special attention should be given and follow-up should be performed to determine whether DNS or PNS occurs, particularly epilepsy and cognitive deficits.

An extract from Taxodium distichum targets hemagglutinin- and neuraminidase-related activities of influenza virus in vitro.

Influenza virus remains an emerging virus and causes pandemics with high levels of fatality. After screening different plant extracts with potential anti-influenza activity, a water extract of Taxodium distichum stems (TDSWex) showed excellent activity against influenza viruses. The EC50 of TDSWex was 0.051 ± 0.024 mg/mL against influenza virus A/WSN/33. TDSWex had excellent antiviral efficacy against various strains of human influenza A and B viruses, particularly oseltamivir-resistant clinical isolates and a swine-origin influenza strain. We observed that the synthesis of viral RNA and protein were inhibited in the presence of TDSWex. The results of the time-of-addition assay suggested that TDSWex inhibited viral entry and budding. In the hemagglutination inhibition assay, TDSWex inhibited the hemagglutination of red blood cells, implying that the extract targeted hemagglutin-related functions such as viral entry. In the attachment and penetration assay, TDSWex showed antiviral activity with EC50s of 0.045 ± 0.026 and 0.012 ± 0.003 mg/mL, respectively. In addition, TDSWex blocked neuraminidase activity. We conclude that TDSWex has bimodal activities against both hemagglutinin and neuraminidase during viral replication.

Inhibition of endosomal fusion activity of influenza virus by Rheum tanguticum (da-huang).

Rhubarb (Rheum tanguticum; da-huang in Chinese medicine) is a herbal medicine that has been used widely for managing fever and removing toxicity. In this study, we investigated how rhubarb inhibits influenza virus during the early stage of the infectious cycle using different functional assays. A non-toxic ethanolic extract of rhubarb (Rex) inhibited several H1N1 subtypes of influenza A viruses in Madin-Darby canine kidney cells, including strains that are clinically resistant to oseltamivir. Time course analysis of Rex addition showed that viral entry was one of the steps that was inhibited by Rex. We also confirmed that Rex effectively inhibited viral attachment and penetration into the host cells. The inhibition of red blood cell haemolysis and cell-cell fusion by Rex suggests that Rex may block haemagglutinin-mediated fusion (virus-endosome fusion) during the fusion/uncoating step. Rex has the capacity to inhibit influenza viruses by blocking viral endocytosis. Thus, rhubarb might provide an alternative therapeutic approach when resistant viruses become more prevalent.

In vitro and in vivo studies of a potent capsid-binding inhibitor of enterovirus 71.

OBJECTIVES: Enterovirus 71 (EV-A71) is an important pathogen that can cause severe neurological symptoms and even death. Our aim was to identify potent anti-EV-A71 compounds and study their underlying mechanisms and in vivo activity. METHODS: We identified a potent imidazolidinone derivative (abbreviated to PR66) as an inhibitor of EV-A71 infection from the screening of compounds and subsequent structure-based modification. Time-course treatments and resistant virus selection of PR66 were employed to study the mode of mechanism of PR66. In vivo activity of PR66 was tested in the ICR strain of new-born mice challenged with EV-A71/4643/MP4. RESULTS: PR66 could impede the uncoating process during viral infection via interaction with capsid protein VP1, as shown by a resistant virus selection assay. Using site-directed mutagenesis, we confirmed that a change from valine to phenylalanine in the 179th amino acid residue of the cDNA-derived resistant virus resulted in resistance to PR66. PR66 increased the virion stability of WT viruses, but not the PR66-resistant mutant, in a particle stability thermal release assay. We further showed that PR66 had excellent anti-EV-A71 activity in an in vivo mouse model of disease, with a dose-dependent increase in survival rate and in protection against virus-induced hind-limb paralysis following oral or intraperitoneal administration. This was associated with reductions of viral titres in brain and muscle tissues. CONCLUSIONS: We demonstrated here for the first time that an imidazolidinone derivative (PR66) could protect against EV-A71-induced neurological symptoms in vivo by suppressing EV-A71 replication. This involved binding to and restricting viral uncoating.

Postcesarean section wound infection caused by Mycobacterium massiliense.

BACKGROUND/PURPOSE: Mycobacterium abscessus subsp. massiliense (a subspecies of the M. abscessus complex) is a rare causative agent of surgical site infection after cesarean section (C section). We tried to seek the common source of infection and unravel the optimal treatment modalities. METHODS: From September 2009 to October 2012, four postpartum women developed C-section wound infections caused by M. massiliense. Speciation of the four isolates was identified using of hsp65, rpoB, and secA1 partial gene sequencing and the Basic Local Alignment Search Tool. The erm(41) and rrl genes were detected for the possibility of inducible macrolide resistance. Pulsed-field gel electrophoresis was used as a tool of molecular epidemiology. All patients underwent intensive intravenous and oral antimycobacterial regimens. Of these patients, three underwent debridement at least once. RESULTS: All four isolates were identified as M. abscessus subsp. massiliense. All of the isolates harbored a truncated erm(41) gene without rrl gene mutations, which explains the susceptibility to clarithromycin and azithromycin. Three isolates were indistinguishable by DNA strain typing, and the fourth strain was clonal with the other three strains. Their infections were not improved in spite of teicoplanin treatment initially. These patients underwent antimycobacterial regimens with/without surgery and were all cured. DISCUSSION: Teicoplanin treatment failure, painful cutaneous nodules, and persistent wound drainage alerted us to the possibility of nontuberculous mycobacterial skin and soft tissue infection. Accurate identification of subspecies, detection of drug resistance genes, susceptibility testing, and optimal antimycobacterial agents with/without surgical debridement are warranted for successful treatment.

Development of ceftriaxone resistance in Salmonella enterica serotype Oranienburg during therapy for bacteremia.

BACKGROUND: The majority of nontyphoid Salmonella infection is identified in children. When an invasive or severe Salmonella infection is encountered, ceftriaxone is recommended for such patients. A 2-year-old girl was hospitalized for the treatment of Salmonella bacteremia and discharged with standard ceftriaxone treatment. She was readmitted to the hospital after 2 days due to the recurrence of the Salmonella bacteremia. The study aimed to unveil the mechanism for the relapse. METHODS: Six isolates (4 blood and 2 stool) were recovered from the patient, with the last two blood isolates being ceftriaxone-resistant. Pulsed-field gel electrophoresis was used for genotyping. Ceftriaxone resistance genes and transferability of the resistance plasmid were examined by molecular methods. RESULTS: All isolates were identified as Salmonella enterica serotype Oranienburg. Five isolates demonstrated almost identical electrophoresis patterns, except that in the two ceftriaxone-resistant isolates an extra band (>100 kb) was noted. A blaCMY-2 gene, carried by a 120-kb conjugative IncI1 plasmid of the sequence type 53, was identified in the two ceftriaxone-resistant isolates. Transfer of the resistance plasmid from one blood isolate to Escherichia coli J53 resulted in the increase of ceftriaxone minimum inhibitory concentration from 0.125 µg/mL to 32 µg/mL in the recipient. CONCLUSION: Ceftriaxone is the standard therapeutic choice for invasive or serious Salmonella infections in children. Pediatricians should be aware of the possibility of resistance development during therapy, especially in areas with a widespread of ceftriaxone resistance genes that are carried by a self-transferrable plasmid, such as the blaCMY-2-carrying IncI1 plasmid identified herein.

Impact of Molecular Epidemiology and Reduced Susceptibility to Glycopeptides and Daptomycin on Outcomes of Patients with Methicillin-Resistant Staphylococcus aureus Bacteremia.

BACKGROUND: Methicillin-resistant Staphylococcus aureus (MRSA) bacteremia was associated with high mortality, but the risk factors associated with mortality remain controversial. METHODS: A retrospective cohort study was designed. All patients with MRSA bacteremia admitted were screened and collected for their clinical presentations and laboratory characteristics. Minimum inhibitory concentration (MIC) and staphylococcal cassette chromosome mec (SCCmec) type of bacterial isolates were determined. Risk factors for mortality were analyzed. RESULTS: Most MRSA isolates from the 189 enrolled patients showed reduced susceptibility to antibiotics, including MIC of vancomycin ≥ 1.5 mg/L (79.9%), teicoplanin ≥ 2 mg/L (86.2%), daptomycin ≥ 0.38 mg/L (73.0%) and linezolid ≥ 1.5 mg/L (64.0%). MRSA with vancomycin MIC ≥ 1.5 mg/L and inappropriate initial therapy were the two most important risk factors for mortality (both P < 0.05; odds ratio = 7.88 and 6.78). Hospital-associated MRSA (HA-MRSA), carrying SCCmec type I, II, or III, was associated with reduced susceptibility to vancomycin, teicoplanin or daptomycin and also with higher attributable mortality (all P < 0.05). Creeping vancomycin MIC was linked to higher MIC of teicoplanin and daptomycin (both P < 0.001), but not linezolid (P = 0.759). CONCLUSIONS: Giving empirical broad-spectrum antibiotics for at least 5 days to treat catheter-related infections, pneumonia, soft tissue infection and other infections was the most important risk factor for acquiring subsequent HA-MRSA infection. Choice of effective anti-MRSA agents for treating MRSA bacteremia should be based on MIC of vancomycin, teicoplanin and daptomycin. Initiation of an effective anti-MRSA agent without elevated MIC in 2 days is crucial for reducing mortality.

Emergence in Taiwan of novel imipenem-resistant Acinetobacter baumannii ST455 causing bloodstream infection in critical patients.

BACKGROUND: Acinetobacter baumannii is one of the most important nosocomial pathogens worldwide. This study aimed to use multilocus sequence typing (MLST) for the epidemiological surveillance of A. baumannii isolates in Taiwan and analyze the clinical presentations and patients' outcome. METHODS: MLST according to both Bartual's PubMLST and Pasteur's MLST schemes was applied to characterize bloodstream imipenem-resistant A. baumannii (IRAB) infection in intensive care units in a medical center. A total of 39 clinical IRAB bloodstream isolates in 2010 were enrolled. We also collected 13 imipenem-susceptible A. baumannii (ISAB) bloodstream isolates and 30 clinical sputum isolates (24 IRAB and 6 ISAB) for comparison. Clinical presentations and outcome of the patients were analyzed. RESULTS: We found that infection by ST455(B)/ST2(P) and inappropriate initial therapy were statistically significant risk factors for mortality. More than one-third of the IRAB isolates belonged to ST455(B)/ST2(P). Most ST455(B)/ST2(P) (80%) carried ISAba1-blaOXA-23, including 10 (66.7%) with Tn2006 (ISAba1-blaOXA-23-ISAba1) in an AbaR4-type resistance island. ST455(B)/ST2(P) appears to evolve from ST208(B)/ST2(P) of clonal complex (CC) 92(B)/CC2(P). In this hospital-based study, A. baumannii ST455 accounted for 38.5% of IRAB bacteremia, with a high mortality of 86.7%. Approximately 85% of ST455(B)/ST2(P)bacteremia had a primary source of ventilation-associated pneumonia. CONCLUSION: We report the emergence in Taiwan of IRAB ST455(B)/ST2(P), which is the current predominant clone of IRAB in our hospital and has been causing bacteremia with high mortality in critical patients.

Pediatric invasive pneumococcal disease in Taiwan following a national catch-up program with the 13-valent pneumococcal conjugate vaccine.

BACKGROUND: Seven-valent pneumococcal conjugate vaccine (PCV) has been available in Taiwan since late 2005. A national catch-up program was launched in Taiwan in 2013, providing 1 dose of 13-valent PCV to children aged 2-5 years. Here, we report the epidemiology of invasive pneumococcal disease (IPD) in children aged ≤5 years in this setting. METHODS: We collected demographic and clinical information for pediatric patients (≤5 years) with IPD between 2008 and 2013. The incidence of IPD was estimated. The logs for PCV import into Taiwan were obtained to evaluate the impact of PCV usage on IPD epidemiology. RESULTS: The overall incidence of IPD in children aged ≤5 years was 15.9 cases per 100,000 person-years. The IPD incidence caused by 7-valent PCV serotypes decreased significantly from 10.0 cases per 100,000 person-years in 2008 to 2.3 cases per 100,000 person-years in 2013. The incidence of IPD caused by serotype 19A increased substantially from 1.7 cases per 100,000 person-years in 2008 to 10.3 cases per 100,000 person-years in 2012, followed by a significant decrease to 5.6 cases per 100,000 person-years in 2013. The significant decrease in the incidence of serotype 19A IPD occurred primarily in children aged 2-5 years. CONCLUSIONS: The 13-valent PCV catch-up program was associated with a significant decrease in serotype 19A IPD incidence in 2013, primarily in children eligible for the 13-valent PCV immunization. Continued surveillance is necessary to assess the further impact of the national catch-up program on pediatric IPD epidemiology in Taiwan.

Fish tank granuloma caused by Mycobacterium marinum.

INTRODUCTION: Mycobacterium marinum causes skin and soft tissue, bone and joint, and rare disseminated infections. In this study, we aimed to investigate the relationship between treatment outcome and antimicrobial susceptibility patterns. A total of 27 patients with M. marinum infections were enrolled. METHODS: Data on clinical characteristics and therapeutic methods were collected and analyzed. We also determined the minimum inhibitory concentrations of 7 antibiotics against 30 isolates from these patients. RESULTS: Twenty-seven patients received antimycobacterial agents with or without surgical debridement. Eighteen patients were cured, 8 failed to respond to treatment, and one was lost to follow-up. The duration of clarithromycin (147 vs. 28; p = 0.0297), and rifampicin (201 vs. 91; p = 0.0266) treatment in the cured patients was longer than that in the others. Surgical debridement was performed in 10 out of the 18 cured patients, and in 1 of another group (p = 0.0417). All the 30 isolates were susceptible to clarithromycin, amikacin, and linezolid; 29 (96.7%) were susceptible to ethambutol; 28 (93.3%) were susceptible to sulfamethoxazole; and 26 (86.7%) were susceptible to rifampicin. However, only 1 (3.3%) isolate was susceptible to doxycycline. DISCUSSION: Early diagnosis of the infection and appropriate antimicrobial therapy with surgical debridement are the mainstays of successful treatment. Clarithromycin and rifampin are supposed to be more effective agents.

Implementation and outcomes of a hospital-wide computerised antimicrobial stewardship programme in a large medical centre in Taiwan.

Antibiotic stewardship is important to address the problem of antimicrobial resistance, but a practical and sustainable strategy to provide stewardship in a large hospital setting is lacking. We developed a hospital-wide computerised antimicrobial approval system (HCAAS) to guide the use of antimicrobial agents in late 2004 in a 3500-bed medical centre in Taiwan. The objective of this study was to evaluate the impacts of HCAAS on the hospital from 2003 to 2009. Following HCAAS deployment, the gradients of consumption over time during the study period of third- and fourth-generation cephalosporins, fluoroquinolones and glycopeptides fell significantly, whilst that of carbapenems increased. The amount and expenditure of antimicrobial use did not increase with the overall healthcare-associated infection rate, and inpatient mortality rate remained stable with a slight decreasing trend. The rate of meticillin-resistant Staphylococcus aureus started to decline in 2002 and continued after HCAAS deployment. There was an increasing isolation of extended-spectrum ß-lactamase-producing Escherichia coli and Klebsiella pneumoniae, presumably leading to the increased use of carbapenems. The isolation rate of Clostridium difficile from patients who developed diarrhoea after antimicrobial therapy did not change over the years, with a mean annual rate of 10.0% after the implementation of HCAAS. HCAAS along with strict infection control measures is necessary to reduce the spread of resistant organisms within the hospital. HCAAS is a sustainable system for providing antibiotic stewardship and exerts a positive impact on the hospital by reducing antimicrobial consumption and expenditure whilst not compromising healthcare quality.

Imipenem heteroresistance induced by imipenem in multidrug-resistant Acinetobacter baumannii: mechanism and clinical implications.

Acinetobacter baumannii has emerged as a major pathogen causing nosocomial infections, particularly in critical patients admitted to the Intensive Care Unit. Increasing resistance to carbapenems in A. baumannii has been observed worldwide. Here we report the clinical impact and mechanism of imipenem heteroresistance (imipenem minimum inhibitory concentration of 6-32 µg/mL with the presence of resistant cells inside the inhibition zone of Etest strips or disks) in multidrug-resistant A. baumannii (MDR-AB). To identify risk factors associated with the emergence of imipenem heteroresistance, a retrospective case-control study was undertaken involving cases with subsequent clinical isolates of the same genotype showing loss of imipenem susceptibility and matched controls with isolates belonging to imipenem-susceptible MDR-AB. The molecular mechanism of heteroresistance was examined. From April 2006 to March 2007, 126 consecutive isolates of MDR-AB were identified from 29 patients. Switch from imipenem susceptibility to heteroresistance was more likely to occur in successive MDR-AB derived from patients who had been exposed to imipenem (length of use 10.9 ± 6.5 days for cases vs. 5.3 ± 4.8 days for controls; P=0.02). An insertion sequence (ISAba1) was found in the promoter region of a class C ß-lactamase gene (bla(ADC-29)) in most imipenem-heteroresistant MDR-AB isolates. In vitro experiments indicated that imipenem heteroresistance, which was associated with overexpression of bla(ADC-29), could be induced by imipenem. Carbapenem use was the only risk factor identified for the emergence of carbapenem-heteroresistant MDR-AB. Physicians should weigh the benefits and risks of each carbapenem-based treatment in managing carbapenem-susceptible MDR-AB infection.

Biofilm production, use of intravascular indwelling catheters and inappropriate antimicrobial therapy as predictors of fatality in Chryseobacterium meningosepticum bacteraemia.

Chryseobacterium meningosepticum usually causes infections in neonates and the immunocompromised. Treatment is handicapped by the organism's inherent multidrug resistance. In this study, the clinical characteristics of patients with C. meningosepticum bloodstream infection (BSI) were retrospectively reviewed. Antimicrobial susceptibilities of the clinical isolates were analysed and their ability to form biofilm was assayed by crystal violet staining and electron microscopy. During 2003-2007, 40 patients with BSI caused by C. meningosepticum were included. Mean patient age was 61.6±22.1 years. Co-morbidities were observed in 38 cases (95.0%) and a high 14-day mortality (52.5%) was observed in these patients. Susceptibility of the isolates was relatively high (>50%) only to piperacillin, piperacillin/tazobactam, trimethoprim/sulfamethoxazole and ciprofloxacin. Multivariate analysis revealed that mortality was associated with the use of central venous catheters, initial inappropriate antimicrobial therapy and higher biofilm production by the organism. Electron microscopy confirmed the formation of biofilm microcolonies on the solid phase of the fibre of nitrocellulose paper in vitro. Time-kill studies showed that biofilm formation helps bacteria to tolerate killing by ciprofloxacin. In conclusion, C. meningosepticum is a biofilm-forming organism. The outcome of patients with biofilm-forming C. meningosepticum infection was adversely affected by the choice of inappropriate antimicrobial therapy and the use of long-term indwelling intravascular catheters.

Pott's puffy tumor in children.

INTRODUCTION: Pott's puffy tumor is characterized by subperiosteal abscess associated with osteomyelitis of frontal bone. Reports are limited for this rare entity in the antibiotics era but increase during past decade. METHODS: We had clinical analysis of a series with six consecutive pediatric patients of Pott's puffy tumor during 20 years in a tertiary medical center via retrospective chart review. One case was described in detail. RESULTS: Male-to-female ratio was 5:1. The mean age at the time of diagnosis was 13 years-3 months. The risk factors were acute sinusitis in two (33%), chronic sinusitis in two (33%), recent head trauma in two (33%), and acupuncture therapy on skull in one (17%). The commonest presenting symptoms were fever, headache, forehead tenderness, vomiting, and fatigue/malaise (100%). Pott's puffy tumor was diagnosed on average the seventh day after fever, and half had intracranial involvement at diagnosis. All had intracranial infections, and most of them had subdural empyema. The most often involved sinus was frontal sinus (100%). The frontal lobe was the most common site of intracranial infection (100%), two thirds of which are polymicrobial from two or more sites. The initial operation was performed on average on the 5.8th days after diagnosis. Half of the patients underwent reoperation. The mortality rate was 17% (one of six). CONCLUSION: The symptoms of Pott's puffy tumor are inconspicuous even though early intracranial involvement often occurred. The importance of early diagnosis and aggravated and prompt treatment with prolonged antibiotic therapy is emphasized for better outcome.

Antibiotic resistance patterns of community-acquired urinary tract infections in children with vesicoureteral reflux receiving prophylactic antibiotic therapy.

OBJECTIVE: The goal was to examine bacterial antimicrobial resistance of recurrent urinary tract infections in children receiving antibiotic prophylaxis because of primary vesicoureteral reflux. METHODS: We reviewed data retrospectively for children with documented vesicoureteral reflux in 2 hospitals during a 5-year follow-up period. The patients were receiving co-trimoxazole, cephalexin, or cefaclor prophylaxis or prophylaxis with a sequence of different antibiotics (alternative monotherapy). Demographic data, degree of vesicoureteral reflux, prophylactic antibiotics prescribed, and antibiotic sensitivity results of first urinary tract infections and breakthrough urinary tract infections were recorded. RESULTS: Three hundred twenty-four patients underwent antibiotic prophylaxis (109 with co-trimoxazole, 100 with cephalexin, 44 with cefaclor, and 71 with alternative monotherapy) in one hospital and 96 children underwent co-trimoxazole prophylaxis in the other hospital. Breakthrough urinary tract infections occurred in patients from both hospitals (20.4% and 25%, respectively). Escherichia coli infection was significantly less common in children receiving antibiotic prophylaxis, compared with their initial episodes of urinary tract infection, at both hospitals. Children receiving cephalosporin prophylaxis were more likely to have an extended-spectrum beta-lactamase-producing organism for breakthrough urinary tract infections, compared with children with co-trimoxazole prophylaxis. Antimicrobial susceptibilities to almost all antibiotics decreased with cephalosporin prophylaxis when recurrent urinary tract infections developed. The extent of decreased susceptibilities was also severe for prophylaxis with a sequence of different antibiotics. However, antimicrobial susceptibilities decreased minimally in co-trimoxazole prophylaxis groups. CONCLUSIONS: Children receiving cephalosporin prophylaxis are more likely to have extended-spectrum beta-lactamase-producing bacteria or multidrug-resistant uropathogens other than E coli for breakthrough urinary tract infections; therefore, these antibiotics are not appropriate for prophylactic use in patients with vesicoureteral reflux. Co-trimoxazole remains the preferred prophylactic agent for vesicoureteral reflux.